[A study on development of enzyme immunoassay of human osteopontin: molecular fragility of serum osteopontin].

Osteopontin, a secreted acidic phosphoglycoprotein with calcium binding activity, has been reported to have diverse functions, including cell adhesion, calcium remodeling, cell signaling and cellular transformation. This recalls us to a concept that the osteopontin can be a marker for certain clinicopathological states. However, a method for analysis of osteopontin has not successfully been developed to be utilized. We have isolated osteopontin from human milk, prepared antibodies specific to it, and tried to establish an enzyme immunoassay (EIA) system to determine osteopontin in serum. It was found that serum osteopontin but not milk osteopontin showed paradoxical phenomena in reaction with the antibodies, i.e., no detectable reaction in immunoelectrophoresis and in EIA, but showing positive reaction in Western blot and in inhibition tests. Taking these phenomena and other circumstantial evidence into account, it was concluded that the serum osteopontin had high molecular fragility, fragmenting into small peptides with paucity of epitopes during incubation time. These findings indicate the possible presence of osteopontins with different molecular characteristics among tissues, and that these characteristics of osteopontin will interfere with the development of quantitative assay.